The multidrug resistance (MDR) gene encodes a plasma membrane glycoprotein 170 KDa (P-gp), which functions as an efflux pump for chemotherapeutic drugs in MDR tumor cells. P-gp is also expressed in normal tissues never exposed to anticancer drugs, predominately in the cells lining the luminal space of a variety of tissues, including the placenta and the endometrium of the gravid uterus. Although the physiological importance of P-gp has not been defined, several functional roles have been proposed. We hypothesized that P-gp in normal tissues may serve as a first line of defense against endogenous xenobiotics in normal cells, regulated by steroids particularly during pregnancy, to protect the developing fetus. Rat placenta and other maternal tissues were examined for developmental regulation of MDR during gestation. The rat ovary expression of mdr1 was initially high and declined after day 6 of gestation. The placenta and uterus had the highest expression of mdr1 at 15 weeks of gestation, followed by 18 weeks of gestation, which is similar to previous reports and mirrors the progesterone profile in the gestating rat. Colon mucosa had a high level of mdr1 expression at 18 weeks, while no detectable differences were observed in the kidney and adrenal throughout gestation. The expression of mdr1 was different from tissue to tissue during early, mid, and late gestation, strongly suggesting that mdr1 may be regulated by gestational hormones in the pregnant rat. The rat SV40-temperature sensitive (ts) A-mutant transformed placental cell line, A950, was used a model for further investigation. The rat trophoblast cell line expressed mdr1b at significantly higher levels in the normal differentiated phenotype (A950 H, nonpermissive temperature) than in A950 L cells (permissive temperature, malignant phenotype). Similar expression of mdr2 in A950 L and A950 H was observed. The high expression of MDR in rat placenta on day 15 coincides with the circulating progesterone levels. To determine if progesterone played a role in the regulation of MDR, both types of A950 cells were treated with two concentrations (5 and 12.5 mu M) of progesterone. Progesterone at 5 mu M significantly increased the expression of MDR in A950 H cells, whereas at 12.5 mu M, progesterone inhibited the expression of MDR. Progesterone at both concentrations had no effect on MDR expression in A950 L cells. Also, we examined dehydroisoandrosterone (DHEA), which is a metabolite of progesterone metabolism in the placenta. DHEA moderately enhanced P-gp production and expression in A950 H cells with no detectable changes in A950 L cells. We demonstrated that MDR was enhanced in differentiated placental cells (A950 H) when exposed to the gestational hormones, progesterone and DHEA.